The research constitutes a two-pronged approach to the understanding of factors attending the development and sustaining of breast cancer. The first deals with metabolic events which may alter the hormonal milieu of the breast. One focus of the metabolic studies will be on fibrocystic disease, a four-fold risk factor for developing breast cancer. We plan to elucidate the origin of the extremely high levels of some steroids in human breast cyst fluid. For example, estriol-3-sulfate (E3-3S) is 100 times more concentrated in cyst fluid than the blood. Facilitated transport against a gradient or synthesis at the cyst lining from androgen precursors followed by preferential secretion of E3-3S into the cyst fluid are two pathways considered. Transport will be studied by infusing 3H-E3-3S into volunteers prior to aspiration of cyst fluid and comparing the tritiated components of the plasma and cyst fluids. Synthesis will be studied by introducing appropriate tritiated androgen precursors into the cyst fluid and determining the metabolites in the cysts aspirated 2 hrs. later. Also, the synthesis of E3-3S from androgen precursors will be studied in homogenates of cyst lining obtained at surgery. Two steroid metabolic pathways manifested by breast tumor in vitro, glucuronide formation and formation of long chain fatty esters will be explored further. We seek to quantify these pathways and seek correlations with parameters such as histopathology, receptor content of the tumor and the course of the disease. The second major thrust of the proposal is on mechanisms of estrogen action in the breast. We have found that the estrogen-stimulated rodent uterus responds early (30 min.) and later (2 hrs.) with induction respectively of a new hydrolase and of plasminogen activator. There is remarkable similarity between the human breast cancer cell line MCF-7 and the rodent uterus with regard to estrogen-stimulated induced proteins. We plan to assay the MCF-7 cultures for the uterine-like proteases, elucidate the biophysical characteristics and assess the consequences in terms of growth and viability of interfering with the elaboration or function of these proteases. These programs will 1) better define the hormonal milieu at the tissue level in an endocrine-related cancer and 2) assess macromolecular events which may relate to the ontogeny of breast cancer.